Eukaryotic Translational and Post translational Gene Regulation

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Characterization of the Drosophila Methoprene -tolerant gene product Juvenile hormone binding and ligand-dependent gene regulation Ken Miura, Masahito Oda, Sumiko Makita and Yasuo Chinzei Department of Medical Zoology, School of Medicine, Mie University, Tsu City, Japan Insect development and reproduction are regulated by two classes of lipid-soluble hormones, the ecdysteroids and juvenile hormones (JHs). The ecdysteroids activate target genes through a heterodimeric receptor complex composing the ecdysone receptor and ultraspiracle (USP) proteins, both of which are members of the nuc- lear steroid ⁄ thyroid ⁄ retinoid receptor superfamily [1]. During insect development, ecdysteroids induce molting while JH determines the nature of each molt by modu- lating the ecdysteroid-induced gene expression cascade [2–4]. In addition, in adult insects, JH has a wide variety of actions related to reproduction, including oogenesis, migratory behaviour and diapause [2,5,6]. The mode of molecular action of JH, however, is still obscure [7]. JHs are a family of esterified sesquiterpe- noids, whose lipid-soluble nature has suggested action directly on the genome through nuclear receptors such as ecdysteroids and the vertebrate steroid ⁄ thyroid ⁄ reti- noid hormones [5,8] although actions of JH through the cell membrane are also documented [9,10]. Many attempts have been made to identify nuclear JH receptors. Jones and Sharp [11] showed that JH III binds to the Drosophila USP protein, which is a homo- logue of the vertebrate retinoid X receptor, promoting Keywords juvenile hormone; juvenile hormone receptor; Methoprene-tolerant; Drosophila; transcription factor Correspondence K. Miura, Department of Medical Zoology, School of Medicine, Mie University, Edobashi 2-174, Tsu514-8507, Japan Fax: +81 59 231 5215 Tel: +81 59 231 5013 E-mail: k-miura@doc.medic.mie-u.ac.jp (Received 27 October 2004, revised 20 December 2004, accepted 4 January 2005) doi:10.1111/j.1742-4658.2005.04552.x Juvenile hormones (JHs) of insects are sesquiterpenoids that regulate a great diversity of processes in development and reproduction. As yet the molecular modes of action of JH are poorly understood. The Methoprene- tolerant (Met) gene of Drosophila melanogaster has been found to be responsible for resistance to a JH analogue (JHA) insecticide, methoprene. Previous studies on Met have implicated its involvement in JH signaling, although direct evidence is lacking. We have now examined the product of Met (MET) in terms of its binding to JH and ligand-dependent gene regu- lation. In vitro synthesized MET directly bound to JH III with high affinity (K d ¼ 5.3 ± 1.5 nm, mean ± SD), consistent with the physiological JH concentration. In transient transfection assays using Drosophila S2 cells the yeast GAL4-DNA binding domain fused to MET exerted JH- or JHA- dependent activation of a reporter gene. Activation of the reporter gene was highly JH- or JHA-specific with the order of effectiveness: JH III  JH II > JH I > methoprene; compounds which are only structur- ally related to JH or JHA did not induce any activation. Localization of MET in the S2 cells was nuclear irrespective of the presence or absence of JH. These results suggest that MET may function as a JH-dependent tran- scription factor. Abbreviations Ahr, aryl hydrocarbon receptor; Arnt, Ahr nuclear translocator; bHLH, basic helix-loop-helix; DBD, DNA binding domain; DCC, dextran-coated charcoal; EGFP, enhanced green Eukaryotic Translational and Post-translational Gene Regulation Eukaryotic Translational and Post-translational Gene Regulation Bởi: OpenStaxCollege After the RNA has been transported to the cytoplasm, it is translated into protein Control of this process is largely dependent on the RNA molecule As previously discussed, the stability of the RNA will have a large impact on its translation into a protein As the stability changes, the amount of time that it is available for translation also changes The Initiation Complex and Translation Rate Like transcription, translation is controlled by proteins that bind and initiate the process In translation, the complex that assembles to start the process is referred to as the initiation complex The first protein to bind to the RNA to initiate translation is the eukaryotic initiation factor-2 (eIF-2) The eIF-2 protein is active when it binds to the high-energy molecule guanosine triphosphate (GTP) GTP provides the energy to start the reaction by giving up a phosphate and becoming guanosine diphosphate (GDP) The eIF-2 protein bound to GTP binds to the small 40S ribosomal subunit When bound, the methionine initiator tRNA associates with the eIF-2/40S ribosome complex, bringing along with it the mRNA to be translated At this point, when the initiator complex is assembled, the GTP is converted into GDP and energy is released The phosphate and the eIF-2 protein are released from the complex and the large 60S ribosomal subunit binds to translate the RNA The binding of eIF-2 to the RNA is controlled by phosphorylation If eIF-2 is phosphorylated, it undergoes a conformational change and cannot bind to GTP Therefore, the initiation complex cannot form properly and translation is impeded ([link]) When eIF-2 remains unphosphorylated, it binds the RNA and actively translates the protein Art Connection 1/4 Eukaryotic Translational and Post-translational Gene Regulation Gene expression can be controlled by factors that bind the translation initiation complex An increase in phosphorylation levels of eIF-2 has been observed in patients with neurodegenerative diseases such as Alzheimer’s, Parkinson’s, and Huntington’s What impact you think this might have on protein synthesis? Chemical Modifications, Protein Activity, and Longevity Proteins can be chemically modified with the addition of groups including methyl, phosphate, acetyl, and ubiquitin groups The addition or removal of these groups from proteins regulates their activity or the length of time they exist in the cell Sometimes these modifications can regulate where a protein is found in the cell—for example, in the nucleus, the cytoplasm, or attached to the plasma membrane Chemical modifications occur in response to external stimuli such as stress, the lack of nutrients, heat, or ultraviolet light exposure These changes can alter epigenetic accessibility, transcription, mRNA stability, or translation—all resulting in changes in expression of various genes This is an efficient way for the cell to rapidly change the levels of specific proteins in response to the environment Because proteins are involved in every stage of gene regulation, the phosphorylation of a protein (depending on the protein that is modified) can alter accessibility to the chromosome, can alter translation (by altering transcription factor binding or function), can change nuclear shuttling (by influencing modifications to the nuclear pore complex), can alter RNA stability (by binding or not binding to the RNA to regulate its stability), can modify translation (increase or decrease), or can change post-translational modifications (add or remove phosphates or other chemical modifications) The addition of an ubiquitin group to a protein marks that protein for degradation Ubiquitin acts like a flag indicating that the protein lifespan is complete These proteins are moved to the proteasome, an organelle that functions to remove proteins, to be degraded ([link]) One way to control gene expression, therefore, is to alter the longevity of the protein 2/4 Eukaryotic Translational and Post-translational Gene Regulation Proteins with ubiquitin tags are marked for degradation within the proteasome Section Summary Changing the status of the RNA or the protein itself can affect the amount of protein, the function of the protein, or how long it is found in the cell To translate the protein, a protein initiator complex must assemble on the RNA Modifications (such as phosphorylation) of proteins in this complex can prevent proper translation from occurring Once a protein has been synthesized, it can be modified (phosphorylated, acetylated, methylated, or ubiquitinated) These post-translational modifications can greatly impact the stability, degradation, or function of the protein Art Connections [link] An increase in phosphorylation levels of eIF-2 has been observed in patients with neurodegenerative diseases such as Alzheimer’s, Parkinson’s, and Huntington’s What impact you ...REVIEW ARTICLE Functional interplay between viral and cellular SR proteins in control of post-transcriptional gene regulation Ming-Chih Lai 1, *, Tsui-Yi Peng 1,2, * and Woan-Yuh Tarn 1 1 Institute of Biomedical Sciences, Academia Sinica, Taipei, Taiwan 2 Institute of Molecular Medicine, National Tsing Hua University, Hsin-Chu, Taiwan Introduction Arginine ⁄ serine (RS) dipeptide repeats are present in a number of cellular proteins, termed SR proteins, that primarily participate in nuclear precursor (pre)-mRNA splicing [1–3]. RS domain variants, such as serine and arginine-rich motifs or arginine–aspartate or arginine– glutamate dipeptide-rich domains, are also found in many nuclear proteins. In addition to the RS domains, SR splicing factors often contain one or more RNA recognition motifs. SR proteins function in both constitutive and regulated splicing via binding to cis-elements of pre-mRNA or interaction with other splicing factors. The RS domain interacts with both proteins and RNAs [1–3]. In particular, intermolecular interactions between SR proteins, which are important for spliceosome assembly and splice site determination during pre-mRNA splicing, are mediated by their RS domains [3]. The RS domain also acts as a nuclear localization signal and targets SR proteins to nuclear speckled domains, where splicing factors are concen- trated, for storage [1]. An important biochemical property of the RS domain is its differential phosphorylation at multiple serine and threonine residues. The RS domain is primarily phos- phorylated by SR protein-specific kinases (SRPKs), and Keywords Alternative splicing; kinases; phosphatases; phosphorylation; post-transcriptional control; pre-mRNA splicing; RS domain; SR proteins; viral problems; virus Correspondence W Y. Tarn, Institute of Biomedical Sciences, Academia Sinica, 128 Academy Road, Section 2, Nankang, Taipei 11529, Taiwan Fax: +886 2 2782 9142 Tel: +886 2 2652 3052 E-mail: wtarn@ibms.sinica.edu.tw *These authors contributed equally to this work (Received 3 November 2008, revised 14 December 2008, accepted 9 January 2009) doi:10.1111/j.1742-4658.2009.06894.x Viruses take advantage of cellular machineries to facilitate their gene expression in the host. SR proteins, a superfamily of cellular precursor mRNA splicing factors, contain a domain consisting of repetitive argi- nine ⁄ serine dipeptides, termed the RS domain. The authentic RS domain or variants can also be found in some virus-encoded proteins. Viral pro- teins may act through their own RS domain or through interaction with cellular SR proteins to facilitate viral gene expression. Numerous lines of evidence indicate that cellular SR proteins are important for regulation of viral RNA splicing and participate in other steps of post-transcriptional viral gene expression control. Moreover, viral infection may alter the expression levels or modify the phosphorylation status of cellular SR proteins and thus perturb cellular precursor mRNA splicing. We review our current understanding of the interplay between virus and host in post-transcriptional regulation of gene expression via RS domain-containing proteins. Abbreviations CTE, constitutive transport element; E4, early region 4; EV, epidermodysplasia verruciformis; HBV, hepatitis B virus; HCV, hepatitis C virus; hnRNP, heterogeneous nuclear ribonucleoprotein; HPV, human papillomavirus; HSV, herpes simplex virus; IRES, internal ribosome entry site; N, nucleocapsid; PP, protein phosphatase; Genome Biology 2006, 7:325 comment reviews reports deposited research interactions information refereed research Meeting report Advances in the genetics and epigenetics of gene regulation and human disease Kristine Kleivi Addresses: Medical Biotechnology, VTT Technical Research Center of Finland, 20521 Turku, Finland and Department of Genetics, Institute for Cancer Research, Rikshospitalet-Radiumhospitalet Medical Center, 0310 Oslo, Norway. Email: kristine.kleivi@vtt.fi Published: 24 August 2006 Genome Biology 2006, 7:325 (doi:10.1186/gb-2006-7-8-325) The electronic version of this article is the complete one and can be found online at http://genomebiology.com/2006/7/8/325 © 2006 BioMed Central Ltd A report on the Human Genome Organisation (HUGO) 11th Human Genome Meeting, Helsinki, Finland, 31 May-3 June 2006. At the recent annual meeting on the human genome in Helsinki, organized by the Human Genome Organisation (HUGO), close to 700 scientists gathered to present and discuss the latest advances in genome research. This report presents some selected highlights. Genome variation, gene expression and disease susceptibility Through their effects on gene expression, polymorphisms in the human genome can contribute to phenotypic variation and disease susceptibility. For many diseases, such as cancer, great effort is being made to study the sequence variants that con- tribute to disease susceptibility. The impact of genetic varia- tion on common diseases was addressed by Kari Stefansson (deCODE Genetics, Reykjavik, Iceland), who gave an update on the identified sequence variants that may increase the risk of developing type 2 diabetes, prostate cancer, myocardial infarction, stroke and schizophrenia. In the past decades, type 2 diabetes has become a major health problem in the Western world, as both its incidence and its prevalence have increased rapidly. Stefansson reported his group’s recent discovery of an inherited variant of the gene TCF7L7, encoding a protein called transcription factor 7-like 2 located on chromosome 10, which is estimated to account for about 20% of the diabetes cases. They have also showed an association between a common genetic variant in the microsatellite DG8S737 at chromosome band 8q24, which may contribute to the development of prostate cancers in European and African populations. Single-nucleotide polymorphism (SNP) genotypes correlated with gene-expression data in breast tumors were presented by Vessela Kristensen (The Norwegian Radium Hospital, Oslo, Norway). For genotyping, she and her colleagues selected sets of genes involved in reactive oxygen species sig- naling (ROS) and the repair of DNA damage caused by ROS - that is, pathways that are generally affected by chemotherapy and radiation therapy. Using various statistical approaches, the genetic association between SNPs in genes involved in the ROS pathways and the expression levels of mRNA transcripts from a panel of breast cancer patients were assessed. Regula- tory SNPs in the genes EGF, IL1A, MAPK8, XPC, SOD2 and ALOX12 were associated with alterations in the expression levels of several transcripts. Kristensen also showed that a set of SNPs were linked to a cluster of transcripts participating in the same functional pathway. Thomas Hudson (McGill University, Montreal, Canada) described several resources and technologies that are avail- able to study the impact of genome variation on gene expres- sion. He and his colleagues systematically studied a subset of genes whose alleles show large differences in expression in lymphoblastoid cell lines. These data were integrated with HapMap data to search for haplotypes associated with mRNA expression at flanking genes. Hudson described the discovery of 16 loci harboring a common haplotype affecting the total expression of a gene, that is, all the alleles of the gene, and of 17 loci that affected relative allelic expression in heterozygous samples. To better understand the mecha- nisms controlling Genome Biology 2008, 9:R177 Open Access 2008Fothet al.Volume 9, Issue 12, Article R177 Research Quantitative protein expression profiling reveals extensive post-transcriptional regulation and post-translational modifications in schizont-stage malaria parasites Bernardo J Foth, Neng Zhang, Sachel Mok, Peter R Preiser and Zbynek Bozdech Address: School of Biological Sciences, Nanyang Technological University, Nanyang Drive, 637551 Singapore. Correspondence: Zbynek Bozdech. Email: zbozdech@ntu.edu.sg © 2008 Foth et al.; licensee BioMed Central Ltd. This is an open access article distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/2.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited. Protein expression profiling in Plasmodium<p>A quantitative time-course analysis of protein abundance for Plasmodium falciparum schizonts using two-dimensional differential gel electrophoresis reveals significant post-transcriptional regulation.</p> Abstract Background: Malaria is a one of the most important infectious diseases and is caused by parasitic protozoa of the genus Plasmodium. Previously, quantitative characterization of the P. falciparum transcriptome demonstrated that the strictly controlled progression of these parasites through their intra-erythrocytic developmental cycle is accompanied by a continuous cascade of gene expression. Although such analyses have proven immensely useful, the correlations between abundance of transcripts and their cognate proteins remain poorly characterized. Results: Here, we present a quantitative time-course analysis of relative protein abundance for schizont-stage parasites (34 to 46 hours after invasion) based on two-dimensional differential gel electrophoresis of protein samples labeled with fluorescent dyes. For this purpose we analyzed parasite samples taken at 4-hour intervals from a tightly synchronized culture and established more than 500 individual protein abundance profiles with high temporal resolution and quantitative reproducibility. Approximately half of all profiles exhibit a significant change in abundance and 12% display an expression peak during the observed 12-hour time interval. Intriguingly, identification of 54 protein spots by mass spectrometry revealed that 58% of the corresponding proteins - including actin-I, enolase, eukaryotic initiation factor (eIF)4A, eIF5A, and several heat shock proteins - are represented by more than one isoform, presumably caused by post-translational modifications, with the various isoforms of a given protein frequently showing different expression patterns. Furthermore, comparisons with transcriptome data generated from the same parasite samples reveal evidence of significant post-transcriptional gene expression regulation. Conclusions: Together, our data indicate that both post-transcriptional and post-translational events are widespread and of presumably great biological significance during the intra-erythrocytic development of P. falciparum. Published: 17 December 2008 Genome Biology 2008, 9:R177 (doi:10.1186/gb-2008-9-12-r177) Received: 19 September 2008 Revised: 1 December 2008 Accepted: 17 December 2008 The electronic version of this article is the complete one and can be found online at http://genomebiology.com/2008/9/12/R177 http://genomebiology.com/2008/9/12/R177 Genome Biology 2008, Volume 9, Issue 12, Article R177 Foth et al. R177.2 Genome Biology 2008, 9:R177 Background Malaria is a serious parasitic disease that causes millions of deaths and incalculable suffering each year. It is caused by unicellular parasites of the genus Plasmodium that are trans- mitted between humans by a mosquito vector. A total of five species of Plasmodium parasites reportedly affect humans [1], with P. falciparum being by far the deadliest. Plasmo- dium parasites are characterized by a complex life cycle, dur- The splicing factor ASF/SF2 is associated with TIA-1-related/ TIA-1-containing ribonucleoproteic complexes and contributes to post-transcriptional repression of gene expression Nathalie Delestienne 1 , Corinne Wauquier 1 , Romuald Soin 1 , Jean-Franc¸ois Dierick 2, *, Cyril Gueydan 1, and Ve ´ ronique Kruys 1, 1 Laboratoire de Biologie Mole ´ culaire du Ge ` ne, Faculte ´ des Sciences, Universite ´ Libre de Bruxelles, Gosselies, Belgium 2 Biovalle ´ e, Proteomics Unit, Charleroi, Belgium Keywords AU-rich elements; hnRNP, heterogenous nuclear ribonucleoprotein; ribonucleoprotein complexes; RNA metabolism; RNA-binding proteins; stress granules Correspondence V. Kruys, Laboratoire de Biologie Mole ´ culaire du Ge ` ne, Institut de Biologie et de Me ´ decine Mole ´ culaires, Universite ´ Libre de Bruxelles, 12 rue des Profs. Jeener et Brachet, 6041 Gosselies, Belgium Fax: +32 2 6509800 Tel: +32 2 6509804 E-mail: vkruys@ulb.ac.be *Present address GSK Biologicals, Wavre, Belgium  These authors contributed equally to this work (Received 10 January 2010, revised 10 March 2010, accepted 25 March 2010) doi:10.1111/j.1742-4658.2010.07664.x TIA-1-related (TIAR) protein is a shuttling RNA-binding protein impli- cated in several steps of RNA metabolism. In the nucleus, TIAR contrib- utes to alternative splicing events, whereas, in the cytoplasm, it acts as a translational repressor on specific transcripts such as adenine and uridine- rich element-containing mRNAs. In addition, TIAR is involved in the general translational arrest observed in cells exposed to environmental stress. This activity is encountered by the ability of TIAR to assemble abortive pre-initiation complexes coalescing into cytoplasmic granules called stress granules. To elucidate these mechanisms of translational repression, we characterized TIAR-containing complexes by tandem affinity purification followed by MS. Amongst the identified proteins, we found the splicing factor ASF ⁄ SF2, which is also present in TIA-1 protein complexes. We show that, although mostly confined in the nuclei of normal cells, ASF ⁄ SF2 migrates into stress granules upon environmental stress. The migration of ASF ⁄ SF2 into stress granules is strictly determined both by its shuttling properties and its RNA-binding capacity. Our data also indi- cate that ASF ⁄ SF2 down-regulates the expression of a reporter mRNA carrying adenine and uridine-rich elements within its 3¢ UTR. Moreover, tethering of ASF ⁄ SF2 to a reporter transcript strongly reduces mRNA translation and stability. These results indicate that ASF ⁄ SF2 and TIA proteins cooperate in the regulation of mRNA metabolism in normal cells and in cells having to overcome environmental stress conditions. In addi- tion, the present study provides new insights into the cytoplasmic function of ASF ⁄ SF2 and highlights mechanisms by which RNA-binding proteins regulate the diverse steps of RNA metabolism by subcellular relocalization upon extracellular stimuli. Structured digital abstract l MINT-7715509: ASF ⁄ SF2 (uniprotkb:Q6PDM2)andTIAR (uniprotkb:P70318) colocalize (MI:0403) by fluorescence microscopy ( MI:0416) Abbreviations ARE, adenine and uridine-rich element; CBB, calmodulin binding buffer; CP, coat protein; FITC, fluorescein isothiocyanate; Fluc, firefly luciferase; HA, haemagglutinin; IP, immunoprecipitation; NLS, nuclear localization signal; NPc, nucleoplasmin core domain; Rluc, Renilla luciferase; RRM, RNA recognition motif; RS, arginine-serine; SG, stress granule; SR, serine-arginine; TAP, tandem affinity purification; TIAR, TIA-1-related. 2496 FEBS Journal 277 Eukaryotic Post-transcriptional Gene Regulation Eukaryotic Posttranscriptional Gene Regulation Bởi: OpenStaxCollege RNA is transcribed, but must be processed into a mature form before translation can begin This processing after an RNA molecule has been transcribed, but before it is translated into a protein, is called ... modification can alter gene expression in many ways Describe how phosphorylation of proteins can alter gene expression 3/4 Eukaryotic Translational and Post -translational Gene Regulation Because... degraded ([link]) One way to control gene expression, therefore, is to alter the longevity of the protein 2/4 Eukaryotic Translational and Post -translational Gene Regulation Proteins with ubiquitin.. .Eukaryotic Translational and Post -translational Gene Regulation Gene expression can be controlled by factors that bind the translation